As methods to determine whether a person suffers from a certain disease or not or to examine the progress of the disease, methods to examine the body non-invasively or methods to analyze or examine biological samples are widely used. For the latter methods, the method to analyze a chemical substance qualitatively and quantitatively as a biochemical marker, which is uniquely generated in the body when he/she suffers from a certain disease, is generally used.
When such a specific disease is a case of a malignant tumor (cancer), the use of polyamine or pterin in human urine or biological fluids as a biochemical marker is suggested to be clinically significant. In addition, the method using natal pteridine, namely oncopterin as a more reliable marker substance is also suggested (See Japanese Published Unexamined Patent Application No. H6-199859, for example). In the present method, urine collected from the patient was hydrolyzed with acid prior to HPLC analysis and oncopterin in the sample is selectively isolated to measure the amount of oncopterin.
In short, the amount of oncopterin in the sample is obtained by (i) running the sample through an anion exchange column, washing and eluting oncopterin with electrolyte solution, (ii) running the eluant through a cation exchange column, washing and eluting oncopterin with acidic solution and isolating oncopterin with a reverse column, if needed, (i) measuring fluorescence intensity derived from the oncopterin in the eluant, and (iv) calculating the amount of oncopterin based on the intensity using the calibration curve.
Then, when the urinary samples collected from a subject with a suspected malignant tumor and a healthy volunteer (a person who does not have suspected diseases such as a malignant tumor, hereinafter the same) are analyzed in this way and the amount of oncopterin in the urinary sample of the subject is significantly higher than that of the healthy volunteer, it is determined that the subject is likely to have a malignant tumor.